Method of administration of platelet-rich plasma to treat an acute cardiac dysfunction

ABSTRACT

Methods are described for using compositions containing platelet-rich plasma for the treatment of acute or chronic dysfunction of cardiac muscle. The method may employ a kit which includes a platelet-rich plasma composition and a pH adjusting agent and a catheter or syringe for delivery of the platelet-rich plasma composition to dysfunctional cardiac muscle. The kit may also include a syringe for withdrawing blood from a patient and a centrifuge device means to obtain platelet-rich plasma.

RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No.11/494,166, filed Jul. 27, 2006 which is a continuation of U.S.application Ser. No. 10/941,124, filed Sep. 15, 2004, now U.S. Pat. No.7,314,617, which is a continuation of U.S. application Ser. No.10/412,821, filed Apr. 11, 2003, now U.S. Pat. No. 6,811,777, whichclaims the priority of Provisional Application No. 60/372,682, filedApr. 13, 2002 which are incorporated herein by reference in theirentirety.

BACKGROUND OF THE INVENTION

1. Field of the Invention

In one embodiment, the present invention is directed to a method for thetreatment of damaged tissue with a composition that includesplatelet-rich plasma. The treated tissue may be connective tissue,cardiac muscle, skeletal muscle, disc material, vertebral bodies,pancreas and other internal organs, brain tissue or spinal cord tissue,or vascular tissue. In a preferred embodiment, the tissue is connectivetissue. The described compositions and methods are also useful in woundhealing and infections.

2. Description of the Related Art

There is a need for treatment protocols for connective tissue injurieswhich are refractory to standard treatments such as anti-inflammatorymedication, bracing, rest and physical therapy. Injuries or other damageto flexible, relatively avascular connective tissues (hereafter“connective tissue” or “connective tissues”) are known to take a verylong time to heal (months or even years). In many cases, injuries toconnective tissues may never heal properly, necessitating surgicalintervention. Connective tissue injuries and disorders have asignificant impact on society. The overall prevalence of these problemsis approximately 140 per 1000 persons in the United States, according toa 1995 survey by the National Center for Health Statistics. The samesurvey estimated the direct costs to be $88.7 billion and the indirectcosts estimated to be up to $111.9 billion for lost productivity.

One example of a connective tissue disorder is lateral epicondylitis.Lateral epicondylitis or “tennis elbow” is a well-known sports medicineand orthopedic disorder. The pathology underlying the disorder isrelated to overuse injury and microtearing of the extensor carpiradialis brevis tendon at the elbow. The body attempts to repair thesemicrotears but the healing process is incomplete in many cases.Pathologic specimens of patients undergoing surgery for chronic lateralepicondylitis reveal a disorganized angiofibroblastic dysplasia. Thisincomplete attempt at repair results in degenerative, immature andavascular tissue. Incompletely repaired tissue is weaker than normaltendon tissue and lacks the strength to function normally. This tissuealso limits the patient by causing pain and negatively impacting thepatient's quality of life.

Similar incomplete repair may be present in other types of connectivetissue injuries or damage, such as patellar tendonitis (Jumper's Knee),Achilles tendonitis (common in runners), and rotator cuff tendonitis(commonly seen in “overhead” athletes such as baseball pitchers),chronic injuries of the ankle ligaments (“ankle sprains”), or ligamenttears.

The pathophysiology of the above-mentioned conditions have been studied.Presently, many different non-operative and operative treatments exist.The non-operative measures include rest, activity modification, oralanti-inflammatory medication and cortisone injections. Rest and activitymodification may help patients with some of these conditions, but thereremains a significant clinical population that are not reachable withthese therapies. Despite widespread use, oral anti-inflammatorymedications have not proven to be useful in controlled studies. Somestudies further suggest that non-steroidal medication may actually havean adverse effect on the healing process for ligament injuries. Also, noacute inflammatory cells have been found in pathologic samples of casesof lateral epicondylitis. Cortisone injections are frankly controversialin the treatment of tendinoses and are contraindicated in acute ligamentinjuries. Several studies note an improvement in patients treated withcortisone in short term follow up. Results beyond one year, however,reveal a high symptom recurrence rate and only an equivocal efficacyrate. These injections also carry the risk of tendon rupture, infection,skin depigmentation, subdermal atrophy and hyperglycemia in diabeticpatients. The operative measures include debridement and repair of theassociated pathologic tendons. However, open or arthroscopic surgery hasmany potential complications such as deep infection, damage toneurovascular structures, and scar formation. The surgery is alsoexpensive and carries the additional risks associated with regional orgeneral anesthesia.

What is needed are compositions and methods for solving the problemsnoted above. Platelet-rich plasma (PRP) is an enrichedplatelet-containing mixture, isolated from whole blood, which isresuspended in a small volume of plasma. While whole blood may containabout 95% red blood cells, about 5% platelets and less than 1% whiteblood cells, PRP may contain 95% platelets with 4% red blood cells and1% white blood cells. PRP can be combined with activating agents such asthrombin or calcium which activate the platelets to release theircontents such as cytokinins and other growth factors. PRP has been usedin medicine, primarily in bone grafting and dental implant applicationsand as part of a composition to use as a surgical adhesive. For example,Landesberg et al (U.S. Pat. No. 6,322,785) disclose an autologousplatelet gel that includes PRP for bone grafts and dental implants. ThePRP is activated by collagen and is applied topically to promote woundhealing.

Antanavich et al. (U.S. Pat. No. 5,585,007) disclose preparation of PRPand use as a tissue sealant. Cochrum (U.S. Pat. No. 5,614,214) disclosesa biopolymer that optionally includes PRP and its use to temporarilyblock arteries and veins. Gordinier et al. (U.S. Pat. No. 5,599,558)disclose a platelet releasate product, which includes platelets bufferedto approximately pH 6.5, for use in a topical application to wounds.

None of the prior art teach treatment of connective tissue with PRP topromote tissue repair. Furthermore, Applicant has discovered that PRPhas even broader application in treating a wide range of tissue injuryand/or damage.

SUMMARY OF THE INVENTION

In one embodiment, the present invention is drawn to a method oftreating an injured tissue in an individual including the steps ofdetermining a site of tissue injury in the individual; and introducing aplatelet-rich plasma composition into and around the site of tissueinjury. In preferred embodiments, the tissue is selected from the groupconsisting of connective tissue, cardiac muscle, skeletal muscle, discmaterial, a vertebral body, brain, spinal cord, and vascular tissue. Ina preferred embodiment, the tissue is a part of an internal organ. In amore preferred embodiment, the internal organ is the pancreas. In aparticularly preferred embodiment, the tissue is a connective tissue.

In preferred embodiments the platelet-rich plasma is titrated to obtaina pH of about 7.3 to 7.5. In a more preferred embodiment, the titrationis performed using a bicarbonate buffer.

In preferred embodiments, the platelet-rich plasma composition includesplatelets obtained from the individual who is to be treated with theplatelet-rich plasma composition. In a preferred embodiment, noexogenous activator is added to the composition prior to itsintroduction into and around the site of injury.

In some embodiments, the method includes the step of mixing into theplatelet composition substantially simultaneously with the introductioninto and around the site of tissue injury, one or more ingredientsincluding but not limited to thrombin, epinephrine, collagen, calciumsalts, and pH adjusting agents.

In one embodiment the present invention is drawn to a platelet-richplasma composition for the treatment of an injured tissue which includesplatelet-rich plasma; and a pH adjusting agent, wherein the compositiondoes not contain an activator of the platelet-rich plasma. In apreferred embodiment, the pH of the platelet-rich plasma composition isadjusted to a pH of about 7.3 to 7.5 with a pH adjusting agent. In amore preferred embodiment, the pH adjusting agent is a bicarbonatebuffer. Preferably, the plasma used for the platelet-rich plasma is froman autologous source.

In one embodiment, the present invention is drawn to a method of makinga platelet-rich plasma composition including the steps of drawing bloodfrom an individual; obtaining a plasma fraction from the blood;isolating platelets from the plasma fraction; resuspending the plateletsin a reduced amount of plasma; and adjusting the pH to provide a pH of7.3 to 7.5 for the resuspended platelets to provide a platelet-richplasma composition, wherein an activator of the platelet-rich plasma isnot added to the platelet-rich plasma composition.

BRIEF DESCRIPTION OF THE DRAWINGS

These and other feature of this invention will now be described withreference to the drawings of preferred embodiments which are intended toillustrate and not to limit the invention.

FIG. 1 shows the average visual analog pain scores at pre-op and 6months post-op.

FIG. 2 shows the average Mayo elbow scores at pre-op and 6 monthspost-op.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

The inventor has surprisingly discovered that certain inventive plateletcompositions are useful for the treatment of several types of tissuedamage or injury. As used herein, the term “injury” is a broad term andis used in the ordinary sense to refer, without limitation, to anytissue damage including a wound, trauma or lesion or any tissuedegeneration. In particular, the inventive platelet compositions may beused to treat incomplete repair of various connective tissues.

In one aspect, the invention relates to a method of treating incompleterepair in a patient's connective tissue comprising: obtaining a plateletcomposition; determining a lesion that comprises the incomplete repairin the patient's connective tissue; and minimally invasively introducingthe platelet composition into and around the lesion. In an aspect, theinvention relates to the method wherein the platelet composition is ator above physiological pH. In an aspect, the invention relates to themethod wherein the platelet composition optionally includes plateletreleasate. In an aspect, the invention relates to the method furthercomprising: mixing into the platelet composition one or more of theingredients selected from thrombin, epinephrine, collagen, calciumsalts, and pH adjusting agents. Also useful are materials to promotedegranulation or preserve platelets, additional growth factors or growthfactor inhibitors, small molecule pharmaceuticals such as NSAIDS,steroids, and anti-infective agents. In an aspect, the invention relatesto the method wherein the patient's connective tissue is selected from:tendons, ligaments, joint capsules, and fascial tissues. In an aspect,the invention relates to the method wherein obtaining the plateletcomposition comprises: drawing blood from a human; and centrifuging theblood to obtain a plasma-rich fraction. In an aspect, the inventionrelates to the method wherein the platelet composition comprisesplatelet-rich plasma. In an aspect, the invention relates to the methodwith the proviso that the platelet composition is substantially freefrom exogenous activators prior to its introduction into and around theregion of the incomplete repair in the patient's connective tissue. Inan aspect, the invention relates to the method wherein the plateletcomposition comprises platelets obtained from the patient.

The invention further relates to a method of treating incomplete repairin a patient's connective tissue comprising: obtaining a plateletcomposition; determining a lesion that comprises the incomplete repairin the patient's connective tissue; and introducing the plateletcomposition into and around the lesion, with the proviso thatsubstantially no activator is added to the platelet composition prior toits introduction into and around the lesion. The invention also relatesto the method wherein the platelet composition is minimally invasivelyintroduced into and around the lesion. The invention also relates to themethod wherein the platelet composition comprises platelet-rich plasma.The invention also relates to the method further comprising: mixing intothe platelet composition substantially simultaneously with its minimallyinvasive introduction into and around the lesion one or more of theingredients selected from thrombin, epinephrine, collagen, calciumsalts, and pH adjusting agents. Also useful are materials to promotedegranulation or preserve platelets, additional growth factors or growthfactor inhibitors, small molecule pharmaceuticals such as NSAIDS,steroids, and anti-infective agents. The invention also relates to themethod wherein the patient's connective tissue is selected from:tendons, ligaments, joint capsules, and fascial tissues. The inventionalso relates to the method wherein introducing the platelet compositioninto and around the lesion comprises activating platelets in theplatelet composition through the action of collagen present in thepatient's connective tissue. The invention also relates to the methodwherein the platelet composition is at or above physiological pH. Theinvention also relates to the method wherein the platelet compositioncomprises platelets obtained from the patient.

In yet another aspect, the invention relates to a compositioncomprising: platelet releasate wherein the composition is at a pHgreater than or equal to physiological pH, and wherein the compositioncomprises substantially no unactivated platelets.

In a further aspect, the invention relates to a method of treatinglesions resulting from acute injury to or chronic disorders of theMedial Collateral Ligament of the knee or elbow, Extensor Carpi RadialisBrevis tendon, Anterior Talofibular Ligament at the ankle, Achillestendon, posterior tibial tendon, patellar tendon, quadriceps tendon,Anterior Cruciate Ligament, Posterior Cruciate Ligament, spinalligaments, disc materials, rotator cuff tendon, or biceps tendonscomprising: obtaining a platelet composition; determining a location ofthe lesion; and introducing the platelet composition into and around thelesion, with the proviso that substantially no activator is added to theplatelet composition prior to its introduction into and around thelesion. In an aspect, the invention relates to the method wherein theplatelet composition is minimally invasively introduced into and aroundthe lesion. In an aspect, the invention relates to the method whereinthe platelet composition comprises platelet-rich plasma. In an aspect,the invention relates to the method further comprising: mixing into theplatelet composition substantially simultaneously with its minimallyinvasive introduction into and around the lesion one or more of theingredients selected from thrombin, epinephrine, collagen, calciumsalts, pH adjusting agents. Also useful are materials to promotedegranulation or preserve platelets, additional growth factors or growthfactor inhibitors, small molecule pharmaceuticals such as NSAIDS,steroids, and anti-infective agents. In an aspect, the invention relatesto the method wherein obtaining the platelet composition comprises:drawing blood from a human; and centrifuging the blood to obtain aplasma-rich fraction. In an aspect, the invention relates to the methodwherein the platelet composition is at or above physiological pH. In anaspect, the invention relates to the method wherein the plateletcomposition comprises platelets obtained from the patient.

In a further aspect, the invention relates to a method of treatinglesions resulting from injury or chronic disorders of cardiac muscle,skeletal muscle, organ systems, vascular tissue, disc material, spinalbodies, spinal cord, and brain tissue comprising: obtaining a plateletcomposition; determining a lesion in need of treatment; and minimallyinvasively introducing the platelet composition into and around thelesion. In an aspect, the invention relates to the method wherein theplatelet composition optionally includes platelet releasate. In anaspect, the invention relates to the method further comprising: mixinginto the platelet composition one or more of the ingredients selectedfrom thrombin, epinephrine, collagen, calcium salts, and pH adjustingagents. Also useful are materials to promote degranulation or preserveplatelets, additional growth factors or growth factor inhibitors, smallmolecule pharmaceuticals such as NSAIDS, steroids, and anti-infectiveagents. In an aspect, the invention relates to the method whereinobtaining the platelet composition comprises: drawing blood from ahuman; and centrifuging the blood to obtain a plasma-rich fraction. Inan aspect, the invention relates to the method wherein the plateletcomposition is at or above physiological pH. In an aspect, the inventionrelates to the method wherein the platelet composition comprisesplatelets obtained from the patient.

In an aspect, the invention relates to the method wherein the patient'sconnective tissue is selected from: tendons, ligaments, joint capsules,and fascial tissues. In an aspect, the invention relates to the methodwherein obtaining the platelet composition comprises: drawing blood froma human; and centrifuging the blood to obtain a plasma-rich fraction. Inan aspect, the invention relates to the method wherein the plateletcomposition comprises platelet-rich plasma. In an aspect, the inventionrelates to the method with the proviso that the platelet composition issubstantially free from exogenous activators prior to its introductioninto and around the region of the incomplete repair in the patient'damaged or injured tissue. In an aspect, the invention relates to themethod wherein the platelet composition comprises platelets obtainedfrom the patient.

The inventive platelet composition may be a biocompatible compositionthat comprises unactivated platelets, activated platelets, plateletreleasate(s), or the like. In an embodiment, the inventive plateletcomposition comprises platelet-rich plasma (PRP).

The term “PRP” as used herein is a broad term which is used in itsordinary sense and is a concentration of platelets greater than theperipheral blood concentration suspended in a solution of plasma, withtypical platelet counts ranging from 500,000 to 1,200,000 per cubicmillimeter, or even more. PRP is formed from the concentration ofplatelets from whole blood, and may be obtained using autologous,allogenic, or pooled sources of platelets and/or plasma. PRP may beformed from a variety of animal sources, including human sources.

Platelets are cytoplasmic portions of marrow megakaryocytes. They haveno nucleus for replication; the expected lifetime of a platelet is somefive to nine days. Platelets are involved in the hemostatic process andrelease several initiators of the coagulation cascade. Platelets alsorelease cytokines involved with initiating wound healing. The cytokinesare stored in alpha granules in platelets. In response to platelet toplatelet aggregation or platelet to connective tissue contact, as wouldbe expected in injury or surgery, the cell membrane of the platelet is“activated” to secrete the contents of the alpha granules. The alphagranules release cytokines via active secretion through the plateletcell membrane as histones and carbohydrate side chains are added to theprotein backbone to form the complete cytokine. Platelet disruption orfragmentation, therefore, does not result in release of the completecytokine.

A wide variety of cytokines are released by activated platelets.Platelet derived growth factor (PDGF), transforming growth factor-beta(TGF-b), platelet-derived angiogenesis factor (PDAF) and plateletderived endothelial cell growth factor (PD-ECGF) and insulin-like growthfactor (IGF) are among the cytokines released by degranulatingplatelets. These cytokines serve a number of different functions in thehealing process, including helping to stimulate cell division at aninjury site. They also work as powerful chemotactic factors formesenchymal cells, monocytes and fibroblasts, among others. For thepurposes of this patent, the term “releasate” refers to the internalcontents of the platelet, including cytokines, which have the potentialto affect another cells' function.

Historically, PRP has been used to form a fibrin tissue adhesive throughactivation of the PRP using thrombin and calcium, as disclosed in U.S.Pat. No. 5,165,938 to Knighton, and U.S. Pat. No. 5,599,558 to Gordinieret al., incorporated in their entirety by reference herein. Activationresults in release of the various cytokines and also creates a clottingreaction within various constituents of the plasma fraction. Theclotting reaction rapidly forms a platelet gel (PG) which can be appliedto various wound surfaces for purposes of hemostasis, sealing, andadhesion.

For instance, PG has been employed to assist with hemostasis in spinesurgery and in a rat liver laceration model. One recent investigationshowed that use of PG resulted in a significant reduction in post-opnarcotic use, a reduction in post-op hemoglobin drop and faster returnof functional range of motion after total knee replacement. PG has alsobeen used in wound sealing. One study employed autologous PG as asealant for craniotomy procedures and noted success in 39 out of 40patients. PG has further been noted to augment bone healing. Onecontrolled randomized prospective trial of 88 patients revealed a 2 foldincrease in radiographic bone maturation and a 50% improvement inhistologic bone density when PG was used (Marx, et al., Oral Surg. OralMed. Oral Path. 1998, vol. 85(6): 638-646). In cardiac and vascularsurgery PG is being employed to reduce the incidence of post-op wounddehiscence and infection (Kjaergard, et al., Eur J Cardio-Thoracic Surg.1996, vol 10: 727-733; Slater, et al., J Ortho Res. 1995, vol 13:655-663; Sumner et al. J. Bone Joint Surg. (Am) 1995, vol 77:1135-1147;Sethi, et al. Presented at International Society for the Study of LumbarSpine, June 2001).

However, in none of the above applications and published documents hasit ever been disclosed that PRP could be used surgically outside of thecontext of PG for any purposes other than platelet transfusions to apatient. It is one unique feature of this invention that platelets donot need to be activated prior to their introduction into a patientduring the practice of the present invention.

In another embodiment, the inventive platelet composition may comprisereleasate from platelets, in addition to platelets themselves. Thereleasate comprises the various cytokines released by degranulatingplatelets upon activation. Many activators of platelets exist; theseinclude calcium ions, thrombin, collagen, epinephrine, and adenosinediphosphate. Releasates according to the invention may be preparedaccording to conventional methods, including those methods described inU.S. Pat. No. 5,165,938 to Knighton, and U.S. Pat. No. 5,599,558 toGordinier et al.

One disadvantage of conventional releasate strategies associated withthe use of PRP as PG is the use of thrombin as a preferred activator. Inparticular, much thrombin used in PG is bovine thrombin, which cancreate problems due to contamination issues regarding Creutzfeldt-Jakobdisease. Many bovine materials are suspect due to possible prioncontamination, and so use of bovine thrombin is disfavored in surgery.Human pooled thrombin is likewise disfavored due to the potential ofcontamination with various materials such as viruses, prions, bacteriaand the like. Recombinant human thrombin might also be used, but isquite expensive.

It is a particular advantage of the present invention that exogenous orextra activators need not be administered to a patient. Collagen, amajor component of connective tissues, is a strong activator ofplatelets. Thus, when the inventive platelet composition is introducedinto and/or around connective tissue, platelets in the plateletcomposition may bind to the collagen and then be activated. This reducesor eliminates the need for administering an exogenous activator such asthrombin. The disadvantages of thrombin use have been noted above. Otherstrong activators, such as calcium ions, can cause severe pain,unintentional clotting, and other undesirable side effects. Thus, in anembodiment of the invention, no or substantially no exogenous activatoris present or added as part of the inventive platelet composition, or isused in the preparation of the inventive platelet composition. Ofcourse, exogenous activators may still be employed if a physiciandetermines that they are medically necessary or desirable.

The platelet composition may be prepared using any conventional methodof isolating platelets from whole blood or platelet-containing bloodfractions. These include centrifugal methods, filtration, affinitycolumns, and the like. If the platelet composition comprises PRP, thenconventional methods of obtaining PRP, such as those disclosed in U.S.Pat. No. 5,585,007 and U.S. Pat. No. 5,788,662 both to Antanavich etal., incorporated herein by reference in their entirety, may beutilized.

The platelet-rich plasma composition may be delivered to an individualin need thereof by convention means which include injection using asyringe or catheter. The platelet rich plasma composition may also bedelivered via a dermal patch, a spray device or in combination with anointment, bone graft or drug. It may further be used as a coating onsuture, stents, screws, plates or some other implantable medical device.Finally, it may be used in conjunction with a bioresorbable drug ordevice.

The site of delivery of the platelet-rich plasma composition is at ornear the site of tissue injury and/or damage. The site of tissue injuryor damage is determined by well-established methods including imagingstudies and patient feedback or a combination thereof. The preferredimaging study used is determined by the tissue type. Commonly usedimaging methods include, but are not limited to MRI, X-ray, CT scan,Positron Emission tomography (PET), Single Photon Emission ComputedTomography (SPECT), Electrical Impedance Tomography (EIT), ElectricalSource Imaging (ESI), Magnetic Source Imaging (MSI), laser opticalimaging and ultrasound techniques. The patient may also assist inlocating the site of tissue injury or damage by pointing out areas ofparticular pain and/or discomfort.

Adjusting the pH of platelet compositions has been used to prolong thestorage time of unactivated platelets, as disclosed in U.S. Pat. No.5,147,776 to Koerner, Jr. and U.S. Pat. No. 5,474,891 to Murphy,incorporated by reference herein. pH may be adjusted using a variety ofpH adjusting agents, which are preferably physiologically toleratedbuffers, but may also include other agents that modify pH includingagents that modify lactic acid production by stored platelets.Especially useful are those pH adjusting agents that result in the pH ofthe platelet composition becoming greater than or equal to physiologicalpH. In an embodiment, the pH adjustment agent comprises sodiumbicarbonate. Physiological pH, for the purposes of this invention, maybe defined as being a pH ranging from about 7.35 to about 7.45. pHadjusting agents useful in the practice of this invention includebicarbonate buffers (such as sodium bicarbonate), calcium gluconate,choline chloride, dextrose (d-glucose),ethylenebis(oxyethylenenitrilo)tetraacetic acid (EGTA),4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES), maleic acid,4-morpholinepropanesulfonic acid (MOPS),1,4-piperazinebis(ethanesulfonic acid) (PIPES), sucrose,N-tris(hydroxymethyl)methyl-2-aminoethanesulfonic acid (TES),tris(hydroxymethyl)aminomethane (TRIS BASE),tris(hydroxymethyl)aminomethane hydrochloride (TRIS.HCl), and urea. In apreferable embodiment, the pH adjusting agent is a bicarbonate buffer,more preferably, sodium bicarbonate.

For the purposes of this patent, the term “tissues” includes but is notlimited to cardiac and skeletal muscle, disc material, vertebral bodies,internal organs, brain and spinal cord tissue, vascular tissue such asarteries and veins and non-differentiated tissue.

For the purposes of this patent, connective tissues comprise tendons,ligaments, fascial tissues, and joint capsules. In a preferableembodiment, connective tissues comprise the Medial Collateral Ligamentof the knee or elbow, Extensor Carpi Radialis Brevis tendon (tenniselbow), Anterior Talofibular Ligament at the ankle, Achilles tendon,Anterior Cruciate Ligament, Posterior Cruciate Ligament, posteriortibial tendon, patellar tendon, quadriceps tendon, rotator cuff tendon,and biceps tendons.

Incomplete repair, as it is used in the context of this patentapplication, may be defined to mean repair that is disorganized,substantially non-existent (such as in the case of an unhealed tear), orotherwise pathological. Disorganized repair is characterized by adisorganized angiofibroblastic dysplasia, with degenerative, immatureand avascular tissue. Such tissue is weaker than normal connectivetissue and lacks the strength to function normally. This tissue alsolimits the patient by causing pain and negatively impacting thepatient's quality of life. Substantially non-existent repair might occurin a situation where a connective tissue is torn and does not healproperly subsequently to the tear. Otherwise pathological repair may beany other type of repair in which the tissue is not repaired to besubstantially the same as the tissue was before tissue repair wasnecessary.

It will be apparent to those skilled in the art that variousmodifications and variations can be made to the present inventionwithout departing from the spirit or scope of the invention. Thus, it isintended that the present invention cover the modifications andvariations of the present invention provided that they come within thescope of the appended claims and their equivalents. The followingexamples are illustrative of the present invention, and are not intendedto limit it.

EXAMPLES Example 1

PRP was prepared using a centrifuge unit made by Harvest (Plymouth,Mass.). (Similar units are available as The Biomet GPS system, the DepuySymphony machine and the Medtronic Magellan machine.) Approximately 55cc of blood was drawn from the patient using a standard sterile syringe,combined with 5 cc of a citrate dextrose solution for anticoagulation,and then spun down to isolate the platelets according to themanufacturer's protocol. These platelets were then resuspended inapproximately 3 cc of plasma. The resulting platelet rich plasmasolution (PRP) was quite acidic and was neutralized with usingapproximately 0.05 cc of an 8.4% sodium bicarbonate buffer per cc of PRPunder sterile conditions to approximately physiologic pH of 7.4. The PRPwas not activated through addition of exogenous activators. This PRPcomposition is referred to herein as autologous platelet extract (APEX).

Example 2

Fifty cc of whole blood is drawn from a patient, and then preparedaccording to the method of Knighton, U.S. Pat. No. 5,165,938, column 3.The PRP is activated according to Knighton using recombinant humanthrombin. The degranulated platelets are spun down and the releasatecontaining supernatant is recovered. The releasate may be optionally pHadjusted to a pH of 7.4 using sodium bicarbonate buffer.

Example 3

Thirty ml of whole blood were drawn from a patient. A plateletcomposition was prepared according to Example 1 of U.S. Pat. No.5,510,102 to Cochrum, incorporated herein by reference in its entirety,except that no alginate is added to the platelet composition.

Example 4

A study of the effects of the PRP compositions described herein was doneusing patients presenting with lateral epicondylitis (tennis elbow)which had failed non-operative treatment (anti-inflammatory medication,bracing rest and physical therapy). The average duration of symptoms ofthese patients was 16.6 months. After obtaining informed consent, thepatients were formally enrolled in the study and randomly assigned toeither the APEX treatment group or control.

The patients were evaluated to obtain a visual pain score, a Mayo ElbowScore, and grip strength. For the visual analog pain score, zero equals“no pain” and 100 equals “the worst pain possible.” The Mayo elbow scoreis an overall function score with a higher score indicating betteroverall function. The values of the two scores were statisticallyevaluated using a paired sample T test with significance set at p<0.05.

Each patient was then given 5 mg of Valium p.o. 30 minutes prior to theinventive tendon procedure. Patients were then prepped and draped in asterile manner. A local anesthetic of 0.5% bupivicaine with epinephrinewas infiltrated into the skin, subcutaneous structures and extensorcarpi radialis brevis tendon. Care was taken to keep the localanesthetic out of the elbow joint.

Two to three minutes after administration of the local anesthetic,approximately 3-5 cc of the APEX composition of Example 1 was introducedinto the extensor carpi radialis brevis tendon at the elbow via a 22gauge needle. The control group received a numbing shot (Bupivicaine).Multiple punctures into the tendon approximately 0.5-1 cm from itsorigin were made. Promptly after the minimally invasive introduction ofeither the APEX composition of Example 1 or the numbing shot, thepatient's arm was immobilized at about 90% of flexion without elevationof the arm or hand. The surgical area was then sterilely dressed and thepatient asked not to move their arm for 30 minutes. Each patients'neurovascular, pain and function status was evaluated shortly after theinjection, and at 30 minutes following the end of the procedure. Eachpatient was given oral narcotic pain medication as needed for the first24-48 hours after the procedure. A formal postoperative stretching andstrengthening program may be initiated at 2-3 days after the procedure.The visual pain score, Mayo Elbow Score, and grip strength were allmonitored post-procedure.

Tables 1 and 2 compare treated patients and control preoperatively andat 8 weeks. 5 patients have been treated with the PRP compositions ofone aspect of the present invention (APEX) while 3 patients received anumbing shot and provided a control. The visual analog pain scores(Table 1) improved an average of 66% for the PRP-treated group versus anaverage improvement of 20.4% in the control group. The MAYO elbow score(Table 2) which tests elbow function improved by an average of 44% inthe PRP-treated group compared to 30.5% in the control group.

TABLE 1 TENNIS ELBOW STUDY. Averaged Visual Analog Pain Scores (0-100)for 5 patients treated with APEX and 3 control patients preoperativelyand at 8 week follow-up. Control APEX (Bupivicaine) Preoperative 77 90Postoperative-8 weeks 26 72 % improvement    66% 20.4% n  5 3

TABLE 2 TENNIS ELBOW STUDY. Averaged Mayo Elbow Scores for 5 patientstreated with APEX and 3 control patients preoperatively and at 8 weekfollow-up. Control APEX (Bupivicaine) Preoperative   57.5 49Postoperative-8 weeks 83 64 % improvement    44% 30.5% n  5 3

The improvement seen in the control may be attributed to the observedphenomenon that insertion of a needle into a painful area without drugdelivery (dry needling), will result in some relief from pain symptoms.However, the significantly larger improvements in elbow function (Table2) and pain level (Table 1) clearly demonstrate the efficacy of the PRPtreatments that are an aspect of the present invention.

Example 5

The patient sample of Example 4 was again evaluated at 6 month timepoints. To date, five patients that are at least six months posttreatment with the APEX composition using the inventive method describedherein have been evaluated. The results are outlined below in Tables 3and 4. Average values for 5 patients tested so far are shown.

TABLE 3 VISUAL ANALOG PAIN SCORE Preoperative Visual Analog Pain ScoreAverage: 77 Six Month Postoperative Visual Analog Pain Score Average: 16Improvement 79%

TABLE 4 MODIFIED MAYO ELBOW SCORE (A measure of overall function)Preoperative Visual Analog Pain Score Average: 57.5 Six MonthPostoperative Mayo Elbow Score Average: 88 Improvement 53%

The Visual Analog Pain Score (Table 3) represents an average of 79%improvement in pain scores at six months. Table 4 represents an averageof 53% improvement in Mayo Elbow score at six months. The data isgraphically depicted in FIG. 1 (visual analog pain scores) and FIG. 2(Mayo elbow score improvement). Importantly, 4 of 5 patients areexcellent (>90 by Mayo elbow score) and one patient is in the goodcategory. All patients were either fair or poor prior to treatment. Evenwith this small sample, the improvement was statistically significant (pvalues=<0.05).

Example 6

A patient presenting with Achilles tendinosis is given 5 mg of Valiump.o. 30 minutes prior to the inventive tendon procedure. The patient isthen prepped and draped in a sterile manner. A local anesthetic of 0.5%bupivicaine with epinephrine is infiltrated into the skin, subcutaneousstructures and Achilles tendon. Care is taken to keep the localanesthetic out of the ankle joint.

Two to three minutes after administration of the local anesthetic,approximately 3-5 cc of the APEX composition of Example 1 is introducedinto the Achilles tendon just above the ankle via a 22 gauge needle.Multiple punctures into the tendon approximately 0.5-1 cm from itsinsertion are made. Promptly after the minimally invasive introductionof the platelet composition of Example 1, the patient's lower leg andfoot are immobilized without elevation of the leg. The surgical area isthen sterilely dressed and the patient is asked not to move their legfor 30 minutes. The patient's neurovascular, pain and function status isevaluated shortly after the injection, and at 30 minutes following theend of the procedure. The patient is given oral narcotic pain medicationas needed for the first 24-48 hours after the procedure. The Achillestendon remains immobilized for one week following the procedure,followed by an formal postoperative stretching and strengthening programinitiated at 8-10 days after the procedure.

Example 7

A patient presenting with a medial collateral ligament tear of the elbowis given 5 mg of Valium p.o. 30 minutes prior to the inventive tendonprocedure. The patient is then prepped and draped in a sterile manner. Alocal anesthetic of 0.5% bupivicaine with epinephrine is infiltratedinto the skin, subcutaneous structures and medial collateral ligament atthe elbow. Care is taken to keep the local anesthetic out of the elbowjoint.

Two to three minutes after administration of the local anesthetic,approximately 3-5 cc of the APEX composition of Example 1 is introducedinto the medial collateral ligament at the elbow via a 22 gauge needle.Multiple punctures into the ligament approximately 0.5-1 cm from itsorigin are made. Promptly after the minimally invasive introduction ofthe platelet composition of Example 1, the patient's elbow and arm areimmobilized at about 90% of flexion without elevation of the arm orhand. The surgical area is then sterilely dressed and the patient isasked not to move their arm for 30 minutes. The patient's neurovascular,pain and function status is evaluated shortly after the injection, andat 30 minutes following the end of the procedure. The patient is givenoral narcotic pain medication as needed for the first 24-48 hours afterthe procedure. An optional formal post-operative stretching andstrengthening program may be initiated at 2-3 days after the procedure.

Example 8 Cardiac Muscle

A patient presents with either an acute (i.e. heart attack) or chronicdysfunction (i.e. congestive heart failure) of cardiac muscle. An APEXcomposition is prepared as described in Example 1 Approximately 0.05 ccof an 8.4% sodium bicarbonate buffer per cc of extract is used to raisethe pH to or slightly above 7.4. The extract is not activated throughthe addition of exogenous agent(s).

The APEX is then introduced into the area of dysfunctional cardiacmuscle via a catheter. The APEX may also be combined with an implantabledevice such as a stent.

Example 9 Skeletal Muscle

A patient presents with weakness or atrophy of skeletal muscle. Thiscould be the result of an injury or after a surgical procedure. Usingthe technique of Example 1, an autologous platelet extract (APEX) isobtained and buffered to physiologic pH.

The area of weakness or atrophy is identified and then after using 0.5%bupivicaine with epinephrine as a local anesthetic, the APEX isintroduced into the muscle via a 22 g needle. This can be done a singletime or it may require multiple injections. Postoperatively, the patientis started on a site specific stretching and strengthening protocol.

Example 10 Disc Material/Vertebral Bodies

A patient presents with low back pain and either bulging or black discson an MRI scan. Using the technique of Example 1, an autologous plateletextract (APEX) is obtained and buffered to physiologic pH.

The disc in question is identified by X-ray guidance and then the APEXis introduced into the disc space via small gauge catheter. Thisprocedure could be done alone or in combination with athermal/radiofrequency ablation procedure. The APEX could also beinjected into a vertebral body that has sustained a compression fracturewith or without the use of a balloon prior to injection.

Example 11 Pancreas/any Internal Organ

A patient presents with diabetes and poor insulin production. Using thetechnique of Example 1, an autologous platelet extract (APEX) isobtained and buffered to physiologic pH.

Under CT guidance and conscious sedation, the APEX is introduced via asmall gauge catheter into the pancreas. The APEX is then injected intothe islet cells to stimulate repair of these cells and thus restoreinsulin production.

Example 12 Brain/Spinal Cord

A patient presents with an acute neurologic deficit such as a spinalcord injury or stroke. Using the technique of Example 1, an autologousplatelet extract (APEX) is obtained and buffered to physiologic pH.

Under MRI guidance and conscious sedation, the APEX is introduced intothe area of injury or deficit. The APEX initiates or assists with repairof the locally damaged cells.

Example 13 Vascular Tissue

A patient presents with an area of hypovascularity in his or her lowerextremities. The patient has a diagnosis of peripheral vascularocclusive disease. Using the technique of Example 1, an autologousplatelet extract (APEX) is obtained and buffered to physiologic pH.

The area of hypovascularity is blocked with bupivicaine and then theAPEX is introduced into either the muscle or soft tissue. The APEXinduces angiogenesis and new blood vessel formation.

Example 14 Wound Healing

A patient presents with a chronic wound that is not healing properly.This could be a diabetic foot ulcer. Using the technique of Example 1,an autologous platelet extract (APEX) is obtained and buffered tophysiologic pH.

The wound is carefully cleansed and debrided as needed. The APEX is thencarefully injected into and around the chronic wound and its edges. Itis held in place with an occlusive bandage or combined with an ointment.This process may be repeated as needed until the wound has healed.

Example 15 Neoplastic Tissue

A patient presents with either a benign or malignant tumor or process.Using the technique of Example 1, an autologous platelet extract (APEX)is obtained and buffered to physiologic pH. The APEX can be used eitherin vivo or in vitro to initiate or induce tumor cell death.

Specifically, the APEX is injected into a solid tumor with CT or MRIguidance via a small catheter. Alternatively, cancer cells after beinggrown in an APEX media could be reintroduced back into the body toattack and kill the remaining tumor. Without intending to be limited bytheory, it is hypothesized that the APEX media has either the ability tocause tumor cell apoptosis (cell death) in vivo or it may have theability to transform cancer cells into normal cells.

Example 16 Infections

A patient presents with a superficial or deep infection. Using thetechnique of Example 1, an autologous platelet extract (APEX) isobtained and buffered to physiologic pH.

The area of the infection is identified and the APEX is then directlyapplied or percutaneously introduced. This could be done under local orgeneral anesthesia and with or without imaging guidance. The APEXtypically has a concentrated number of white blood cells along withplatelets. This combination of white blood cells and platelets eithercontrols or eliminates an infection.

Example 17 Cell Cultures of Any Tissue

A researcher or clinician wishes to grow a cell culture of eitherfibroblasts or osteoarthritic cartilage cells. Using the technique ofExample 1, an autologous platelet extract (APEX) is obtained andbuffered to physiologic pH.

The cells are then isolated and grown in a media rich in the APEX invarious conditions and dilutions. The APEX promotes cell differentiationand production of proteins such as collagen. The APEX may augment orpromote the ability of the cells to transform into normal cells. Withoutintending to be limited by theory, it is hypothesized the APEX mayconvert the osteoarthritic cartilage cells to a more functional cellline that is reinjected into a diseased or injured joint. Alternatively,the APEX is directly introduced into an osteoarthritic joint to reversethe course of the disease. This is done under local anesthesia in asterile manner.

Finally, the APEX may be used to help grow and differentiate any tissueor cell line in vivo or in vitro.

It will be understood by those of skill in the art that numerous andvarious modifications can be made without departing from the spirit ofthe present invention. Therefore, it should be clearly understood thatthe forms of the present invention are illustrative only and are notintended to limit the scope of the present invention.

What is claimed is:
 1. A method of treating myocardial infarction in anindividual comprising introducing an unactivated platelet-rich plasmacomposition via catheter or syringe into an area of dysfunctionalcardiac muscle using a kit comprising: a syringe for withdrawing bloodfrom a patient; a centrifuge device to obtain platelet-rich plasma; a pHadjusting agent for the platelet-rich plasma to provide a platelet-richcomposition having a pH of about 7.3 to 7.5; and a catheter or syringefor delivery of the platelet-rich plasma composition to dysfunctionalcardiac muscle, thereby treating myocardial infarction.
 2. The method ofclaim 1, wherein the pH adjusting agent is a bicarbonate buffer.
 3. Themethod of claim 1, wherein the introduction of the platelet-rich plasmais combined with an implantable device.
 4. The method of claim 3,wherein the implantable device is a stent.
 5. The method of claim 1,wherein the platelet-rich plasma composition comprises plateletsobtained from the individual.